本文關(guān)鍵詞：飛秒激光顯微成像及CARS光譜探測(cè)研究　出處：《南昌航空大學(xué)》2015年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 飛秒激光 雙光子熒光 顯微成像 CARS光譜

【摘要】：隨著科技的發(fā)展,人類希望觀察到一個(gè)更加微觀的世界,不僅研究生物組織或者細(xì)胞的結(jié)構(gòu)形態(tài),還要了解生物最基本的組成成分——分子或者原子。近年來,與此相關(guān)技術(shù)取得了很大成就,如對(duì)生物組織或者細(xì)胞的顯微成像和物質(zhì)分子檢測(cè)等,其中飛秒激光雙光子熒光顯微成像技術(shù)和相干反斯托克斯拉曼散射(CARS)光譜探測(cè)技術(shù)作為新興的生物組織顯微成像和分子結(jié)構(gòu)檢測(cè)的重要手段,以其獨(dú)特的優(yōu)勢(shì)吸引了很多研究著的研究。飛秒激光雙光子熒光顯微成像技術(shù)作為生物組織和細(xì)胞顯微成像的有效手段之一,其激發(fā)光源選用了近紅外的飛秒激光,不僅雙光子熒光波長(zhǎng)遠(yuǎn)離了激發(fā)波長(zhǎng),實(shí)現(xiàn)了暗場(chǎng)成像大大提高了信噪比,還對(duì)生物樣品的光損傷小、穿透能力強(qiáng)、可以獲得較大成像深度,相對(duì)于單光子熒光共焦顯微成像這是其重要的優(yōu)點(diǎn)。本文理論上推導(dǎo)了雙光子的激勵(lì)過程,分析了雙光子熒光空間分布強(qiáng)度,繼而相比于單光子共焦顯微成像分析其優(yōu)勢(shì)。在理論研究基礎(chǔ)上,利用鎖模鈦寶石激光器搭建了一套飛秒激光雙光子熒光顯微成像系統(tǒng),自行編寫了系統(tǒng)控制軟件,設(shè)計(jì)了信號(hào)掃描采集系統(tǒng)。本文通過實(shí)驗(yàn)測(cè)量了Rhodamine B溶液在不同濃度下的雙光子熒光光譜,總結(jié)了雙光子熒光光譜在Rhodamine B溶液濃度不斷增大的情況下,熒光光譜特征峰產(chǎn)生的非線性紅移規(guī)律,其中心波長(zhǎng)隨著Rhodamine B溶液濃度增大向最大熒光發(fā)射波長(zhǎng)610nm慢慢靠近。通過一系列實(shí)驗(yàn),研究了樣品與物鏡焦點(diǎn)之間距離對(duì)成像效果的影響,印證了理論推導(dǎo)結(jié)果,即只有在焦點(diǎn)附近才能產(chǎn)生雙光子熒光過程。本文對(duì)Rhodamine B形成的微結(jié)構(gòu)樣品進(jìn)行了顯微成像,并就其成像結(jié)果進(jìn)行了分析研究。本文基于相干反斯托克斯拉曼散射(coherent Anti-stokes Raman scattering,CARS)技術(shù)對(duì)分子結(jié)構(gòu)檢查進(jìn)行了研究。在理論研究了CARS的三階非線性過程之后,采用時(shí)間分辨方法抑制CARS光譜信號(hào)中非共振背景噪聲。為了獲取比較完整分子振動(dòng)模式對(duì)應(yīng)的CARS光譜,利用飛秒脈沖抽運(yùn)光子晶體光纖(Photonic Crystal Fiber,PCF)產(chǎn)生超連續(xù)譜激光作為激勵(lì)光源,并以此搭建了超連續(xù)譜激發(fā)時(shí)間分辨CARS系統(tǒng)。在此系統(tǒng)中,鎖模鈦寶石激光器輸出的飛秒脈沖一分為二,其中一部分飛秒脈沖抽運(yùn)光子晶體光纖產(chǎn)生超連續(xù)譜激光,作為CARS過程的泵浦光和斯托克斯光;另一部分飛秒脈沖通過適當(dāng)?shù)臅r(shí)間延遲作為CARS過程的探針光。本文通過實(shí)驗(yàn)測(cè)量了二甲基亞砜分子振動(dòng)范圍在500cm-1-3500 cm-1內(nèi)的CARS光譜信息,并討論了溫度對(duì)CARS光譜信號(hào)的影響。
[Abstract]:With the development of science and technology, human beings want to observe a more microscopic world, not only to study the structure and morphology of biological tissues or cells, but also to understand the most basic components of biology-molecules or atoms. Great achievements have been made in related techniques, such as microscopic imaging of biological tissues or cells and detection of substances and molecules. Among them, femtosecond laser two-photon fluorescence microscopy and coherent anti-Stokes Raman scattering (CARSs) spectrum detection techniques are important means of tissue microscopic imaging and molecular structure detection. The femtosecond laser two-photon fluorescence microscopy is one of the effective methods for tissue and cell microimaging. The excitation light source is femtosecond laser with near infrared. The two-photon fluorescence wavelength is far away from the excitation wavelength, and the dark field imaging greatly improves the SNR, and the light damage to biological samples is small, and the penetration ability is strong. Compared with single photon fluorescence confocal microscopy, the imaging depth can be obtained, which is an important advantage. In this paper, the two-photon excitation process is derived theoretically, and the intensity of two-photon fluorescence spatial distribution is analyzed. On the basis of theoretical research, a femtosecond laser two-photon fluorescence microscopy imaging system is built by using mode-locked Ti: sapphire laser. The system control software was written and the signal scanning acquisition system was designed. The two-photon fluorescence spectra of Rhodamine B solution at different concentrations were measured experimentally. In this paper, the nonlinear redshift of the characteristic peaks of two-photon fluorescence spectra with the increasing concentration of Rhodamine B solution is summarized. The center wavelength is approaching to the maximum fluorescence emission wavelength of 610nm with the increase of Rhodamine B solution concentration. A series of experiments have been carried out. The influence of the distance between the sample and the focus of the objective lens on the imaging effect is studied, and the theoretical results are verified. That is, only near the focal point can produce the two-photon fluorescence process. In this paper, the microstructure of Rhodamine B formed by microscopic imaging. The imaging results are analyzed and studied. This paper is based on coherent anti-Stokes Anti-stokes Raman scattering. After studying the third-order nonlinear process of CARS in theory. The time-resolved method is used to suppress the non-resonant background noise of the CARS spectral signal. In order to obtain the CARS spectrum corresponding to the complete molecular vibration mode. Photonic Crystal fiber PCFs are generated by femtosecond pulse pumped photonic crystal fiber (Photonic Crystal fiber) as excitation light source. In this system, the femtosecond pulse output from the mode-locked Ti: sapphire laser is divided into two parts. One part of femtosecond pulse pumped photonic crystal fiber produces supercontinuum spectrum laser, which is used as pump light and Stokes light in CARS process. Another part of femtosecond pulse is used as probe light of CARS process by proper time delay. The vibrational range of dimethyl sulfoxide molecule is measured in 500cm-1-3500 experimentally. CARS spectral information in cm-1. The influence of temperature on CARS spectral signal is discussed.
【學(xué)位授予單位】：南昌航空大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：TN249

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